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Biomedical subjects

J J Robinson

Publications and source records attributed to J J Robinson.

At least 19 recordsLinked to original sources
PubMed1996

Effect of continuous infusion of thyrotrophin-releasing hormone on plasma prolactin and on ovarian activity in melatonin-treated ewes.

Twelve anoestrous ewes maintained under natural photoperiod at 57 degrees N received an oral dose of 3 mg melatonin daily at 15:00 h from 1 May. Starting 41 days later and extending from 11 June until 5 September, six of the ewes were also infused continuously with 0.8 mg thyrotrophin-releasing hormone (TRH) day-1 via subcutaneous osmotic minipumps. The remaining six ewes acted as controls. Behavioural oestrus, ovulation rate and luteal function were determined by exposure to a vasectomized ram, laparoscopy and the measurement of progesterone in peripheral plasma, respectively. TRH infusion stimulated a sustained increase (P < 0.001) in plasma concentrations of thyroxine, tri-iodothyronine and prolactin (thyroxine: 158 +/- 9.3 and 65 +/- 7.7 nmol l-1 for TRH-infused and control ewes, respectively; tri-iodothyronine, 2.6 +/- 0.12 and 1.1 +/- 0.19 nmol l-1 and prolactin, 57 +/- 12 and 11 +/- 2 micrograms l-1). No ewes were in oestrus before TRH infusion and the mean number of behavioural oestrous cycles per ewe during the infusion period was 1.3 +/- 0.33 and 2.5 +/- 0.34 for TRH-infused and control ewes, respectively (P < 0.05). Corresponding mean intervals from 1 May to the onset of the first luteal phase (progesterone > 1 ng ml-1) were 88 +/- 8.9 and 79 +/- 3.5 days (not significant). TRH infusion had no effect on the mean numbers of corpora lutea (1.7 +/- 0.14 and 1.6 +/- 0.20 for TRH-infused and control ewes, respectively), but was associated with a lower mean incidence of normal luteal phases (1.5 +/- 0.43 versus 2.7 +/- 0.21, P = 0.052). Abnormalities in luteal function included delayed initial expression, extended ovarian cycles, suprabasal periovulatory progesterone concentrations and protracted periods of low progesterone secretion between successive ovarian cycles. Thus continuous TRH infusion suppressed plasma prolactin, doubled the circulating concentrations of thyroxine and tri-iodothyronine, and was associated with a wide range of abnormalities in ovarian function and endocrine status, the nature of which varied between ewes.

Animals↗
PubMed1996

Excess dietary urea intake in ewes and its effect on ovulation rate and embryo development.

The effect of excess dietary urea on ovulation and early embryo development of sheep was studied. Thirty Border Leicester x Scottish Black face ewes randomly assigned to three treatments were given a basal control diet (C) which met energy requirements for body weight maintenance. Other treatments were basal diet plus 24 g of urea/day (low urea, L) or plus 48 g (high urea, H)/day. The reproductive cycles of the ewes were synchronized using a single injection of prostaglandin (PGF2 alpha) and progesterone by an intravaginal controlled internal drug release (CIDR) device for 12 days. Ovulation was induced by the use of pregnant mare serum gonadotrophin (PMSG). Ewes were inseminated approximately 52 hours after CIDR device removal using a laparoscopic technique. Embryos were recovered at Day 4 or Day 11 after insemination from half of the ewes from each treatment group. There were no significant differences in ovulation rates among the three groups. The embryo recovery rates were not affected by day of recovery. At embryo recovery on Day 4, 7/13 in C, 3/6 in L and 0/7 in H embryos were morulae. After 72 hours of in vitro culture 6/10 in C, 2/3 in L and 0/4 in H embryos developed to the blastocyst stage. Pregnancies sustained were C 6/8, L 5/7 and H 1/3 of the autotransfers. Throughout the experiment plasma urea levels were significantly affected by diet (p < 0.01). Plasma ammonia levels in the H group were significantly higher than those in the C and L groups (p < 0.05) for 4 hours after each feed. There was no treatment effect on plasma progesterone concentration. The luteinizing hormone (LH) surge onset time and amplitude were not correlated to ovulation rate and were not affected by treatment. It is concluded that high circulating concentrations of plasma urea and ammonia have an adverse effect on early embryo development. This effect was independent of any alterations in progesterone and LH concentrations.

Ammonia↗
PubMed1995

Identification of a component of the sea urchin hyaline layer, HLC-175, which undergoes proteolytic processing during development.

To define the role(s) played by the sea urchin extraembryonic matrix, the hyaline layer, we have previously purified and characterized a number of the protein components of this structure. We are currently studying the timing and significance of the proteolytic processing of these species. The localization of HLC-175 in the egg and 1-hr-old embryo was determined by indirect immunofluorescence analysis. The relationship between HLC-175 and the 109- and 81 kDa species was determined by a combination of native gel electrophoresis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) under non-reducing conditions and protein gel blot analyses using the anti-175, -109 and 81 kDa antisera. Using gel exclusion chromatography we have fractionated a mixture of proteins extracted from the surface of 1-hr-old sea urchin embryos. A set of fractions eluting from the column contained three species of apparent molecular masses 175-, 109- and 81 K. These species comigrated on analysis by either non-reducing SDS-PAGE or native gel electrophoresis. Inclusion of the reducing agent, dithiothreitol, in the solubilizing solutions abolished comigration of these polypeptides. When polyclonal antisera were prepared against each of these antigens cross-reactivity between the 175- and 109 kDa species and between the 175- and 81 kDa species was detected. Developmental protein gel blot analyses revealed a precursor-product relationship between the 175- and the 109- and 81 kDa polypeptides. Indirect immunofluorescence analysis confirmed the localization of HLC-175 to the hyaline layer. The results reported here clearly identify HLC-175 as a component of the hyaline layer.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

The management of diabetes in adolescents and young adults: a preliminary case study.

This paper describes one part of a first-stage study concerned with the care received by a group of adolescents and young adults with insulin-dependent diabetes mellitus (IDDM) in one district health authority. The paper reports findings from a self-selected sample of 30 patients aged between 15 and 25. Each of the 30 patients was interviewed during a routine appointment at a diabetes clinic. The patients' doctor at the clinic and one of the diabetes specialist nurses also filled in a short questionnaire for each of the patients interviewed. The main finding was that although all 30 patients interviewed reported that their diabetes control was at least moderately good, a significant number of patients had blood glucose levels above the recommended norm. The study lends support to the widely held belief that young adults with diabetes have particular difficulties in maintaining blood glucose levels within a normal range. It is suggested that processes within the current mode of care may be contributing to this poor level of control. Recommendations for a controlled trial of specialist nurse versus doctor care are made.

Adolescent↗
PubMed1994

Cloning and characterization of HLC-32, a 32-kDa protein component of the sea urchin extraembryonic matrix, the hyaline layer.

A 32-kDa protein (HLC-32) was purified from sea urchin embryos and characterized. In indirect immunofluorescence analysis this species was found to be distributed throughout the cytoplasm of the unfertilized egg but within 1 hr following fertilization was localized to the hyaline layer with some residual immunofluorescence remaining within the cytoplasm of the single cell embryo. By the early blastula stage the antigen was also detected in the basal lamina, indicating a bidirectional transport mode for this protein. A cDNA clone was isolated from a lambda Zap expression library. Sequence analysis revealed a protein rich in acidic (12.4 mole%) and basic (14.3 mole%) residues. Comparative sequence analysis revealed 41 and 47% amino acid sequence homology with two butanol-extractable proteins previously isolated from blastula stage embryos of the sea urchin Paracentrotus lividus. DNA gel blot analysis indicated that the gene encoding HLC-32 belonged to a small multigenic family, while in RNA gel blot analyses a 1.4-kB transcript was detected in ovaries but not gut, testes, mature eggs, or various stage embryos. At the early blastula stage of development this protein underwent a post-translational modification reaction which resulted in the loss of a 4-amino-acid peptide fragment from the amino-terminal end of this species.

Amino Acid Sequence↗
PubMed1994

Phospholipase D-dependent and -independent activation of the neutrophil NADPH oxidase.

Stimulation of the respiratory burst of human neutrophils by fMet-Leu-Phe (in the absence of cytochalasin B) is largely unaffected when the activities of protein kinase C and phospholipase D are inhibited. This has been confirmed using three separate assays to measure the respiratory burst. However, whilst these enzymes are not required for the initiation or maximal rate of oxidant generation, they are required to sustain oxidase activity. In contrast, in the presence of cytochalasin B, fMet-Leu-Phe stimulated oxidase activity is much more dependent on phospholipase D activity. It is proposed that (in the absence of cytochalasin B) activation of the NADPH oxidase utilises cytochrome b molecules that are already present on the plasma membrane and activation occurs independently of phospholipase D and protein kinase C. Once these complexes are inactivated, then new cytochrome b molecules must be recruited from sub-cellular stores. This translocation and/or activation of these molecules is phospholipase D dependent. Some support for this model comes from the finding that the translocation of CD11b (which co-localises with cytochrome b) onto the cell surface is phospholipase D dependent.

1-Butanol↗
PubMed1994

Stimulation of reactive oxidant production in neutrophils by soluble and insoluble immune complexes occurs via different receptors/signal transduction systems.

Cell-free synovial fluid from patients with rheumatoid arthritis contains soluble and insoluble IgG-containing immune complexes which activate reactive oxidant production in human neutrophils. In this report we have measured the effects of inhibitors of signal transduction pathways on neutrophil activation by these complexes and also following activation by synthetic soluble and insoluble immune complexes made from human serum albumin (HSA) and anti-(HSA) antibodies. In all aspects studied, the soluble rheumatoid complexes and the soluble synthetic complexes were indistinguishable in the ways in which they activated neutrophils. Activation of reactive oxidant production in response to these soluble complexes was completely inhibited by pertussis toxin (indicating G-protein coupling of receptor occupancy), completely insensitive to staurosporine (indicating that oxidant production did not require protein kinase C activity), only marginally (< 30%) inhibited by butanol (indicating that dependence upon activity of phospholipase D was minimal), and completely inhibited by chloracysine, an inhibitor of phospholipase A2. In contrast, activation of reactive oxidant production in response to the insoluble rheumatoid or insoluble synthetic immune complexes was largely pertussis toxin insensitive, inhibited by > 50% by staurosporine, inhibited by > 50% by butanol, and completely inhibited by chloracysine. These results show that the receptor-mediated signal transduction systems activated by the soluble and insoluble immune complexes are different. Because the soluble complexes activate a transient burst of reactive oxidant secretion from primed neutrophils, the mechanisms regulating either the release or the intracellular production of oxidants within rheumatoid joints are distinct and hence may be pharmacologically modified independently of each other.

Alkaloids↗
PubMed1994

Role of Fc gamma receptors in the activation of neutrophils by soluble and insoluble immunoglobulin aggregates isolated from the synovial fluid of patients with rheumatoid arthritis.

OBJECTIVES: Synovial fluid from patients with rheumatoid arthritis contains both soluble and insoluble immunoglobulin aggregates which activate reactive oxidant production in human neutrophils. The objectives were to determine the roles played by Fc gamma receptors in activation of neutrophils by these complexes. METHODS: Pronase treatment was used to remove Fc gamma RIII from the neutrophil surface and blocking monoclonal antibodies were used to prevent the binding of complexes to Fc gamma RII and Fc gamma RIII. RESULTS: When Fc gamma RIII was removed from the cell surface by pronase treatment, activation by the soluble aggregates did not occur [mean (SD) inhibition 89 (16)%, n = 6] whereas activation via the insoluble aggregates was less affected [34 (16)%, n = 6]. Blocking the binding to Fc gamma RIII with antibodies decreased activation in response to the soluble aggregates [mean (SD) inhibition 71 (22)%, n = 8] but again had a lower effect on activation by the insoluble aggregates [40 (17)%, n = 9]. When binding to Fc gamma RII was blocked, activation via the soluble aggregates was substantially inhibited [mean (SD) 93 (13)%, n = 8] whereas that via the insoluble aggregates was inhibited to a much lesser extent [28 (38)%, n = 9]. When Fc gamma RII and III were simultaneously blocked, activation by the insoluble aggregates was only inhibited by 45% [(19), n = 5]. CONCLUSION: These data thus indicate that activation of human neutrophils by soluble immunoglobulin aggregates from rheumatoid synovial fluid occurs via cooperative occupancy of both Fc gamma RII and III: perturbation of binding to either of these receptor classes will abrogate activation.

Antibodies, Monoclonal↗
PubMed1993

Activation of neutrophils by soluble and insoluble immunoglobulin aggregates from synovial fluid of patients with rheumatoid arthritis.

OBJECTIVES: Previous work has shown that synovial fluid isolated from patients with active rheumatoid arthritis contains soluble (not sedimented by centrifugation at 11,600 g for two minutes) and insoluble (sedimented by centrifugation at 11,600 g for two minutes) immunoglobulin aggregates that are capable of activating reactive oxidant production by bloodstream neutrophils. The purpose of this study was to determine which of these types of immunoglobulin aggregates activated the secretion of reactive oxygen metabolites and granule enzymes from neutrophils. METHODS: Cell free synovial fluid (from patients with rheumatoid arthritis) was added to neutrophils isolated from blood of healthy controls that had been incubated in the presence and absence of granulocyte-macrophage colony stimulating factor (GM-CSF). Reactive oxidant production was measured by luminol chemiluminescence (which detects both intracellular and extracellular oxidant production) and by cytochrome c reduction (which measures superoxide secretion). RESULTS: The soluble aggregates only activated neutrophils that were previously primed, and activated a rapid and transient burst of reactive oxidant secretion. On the other hand, the insoluble aggregates activated primed and unprimed neutrophils with similar efficacy and most of the oxidants generated (especially in unprimed cells) were intracellular. The soluble aggregates, but not the insoluble aggregates, also activated the secretion of myeloperoxidase from neutrophils that had either been pretreated with cytochalasin B or primed with GM-CSF. CONCLUSION: It is thus proposed that these soluble immunoglobulin aggregates are responsible for activation of the release of tissue damaging granule enzymes and reactive oxidants from primed neutrophils within the rheumatoid joint.

Adult↗
PubMed1993

Receptor expression in synovial fluid neutrophils from patients with rheumatoid arthritis.

OBJECTIVES: The aim of this study was to determine if neutrophils isolated from the blood and synovial fluid of patients with rheumatoid arthritis had patterns of receptor expression resembling those of blood neutrophils from controls which had been activated and primed in vitro. METHODS: Fluorescence activated cell sorting was used to measure receptor expression in paired blood and synovial fluid neutrophils from patients and in control neutrophils exposed to phorbol myristate acetate and granulocyte-macrophage colony stimulating factor. RESULTS: There was no significant difference in the patterns of receptor expression in blood neutrophils from patients and healthy controls, but neutrophils in the synovial fluid had been primed and activated within the joint. About 50% of rheumatoid synovial fluid neutrophil samples expressed Fc gamma RI, a high affinity receptor for monomeric IgG, which is only expressed in neutrophils exposed to cytokines. CONCLUSIONS: Synovial fluid neutrophils are activated and primed within the inflamed joint and hence their ability to respond to activating factors such as immune complexes will be modulated. As the expression of Fc gamma RI requires active biosynthesis, this work indicates that selective gene activation occurs when neutrophils are recruited into rheumatoid joints.

Antigens, CD↗
PubMed1993

Effect of chronic treatment with a GnRH agonist to suppress pulsatile LH secretion on the ability of exogenous melatonin to advance oestrous cyclicity in ewes.

Thirty-two Scottish Blackface ewes that lambed outdoors in March and were weaned at the end of April and individually penned indoors under the natural photoperiod at 57 degrees N were used to determine whether the ovine ovary that was deprived of gonadotrophic support was capable of early activation by melatonin. From 5 May (day 0), 16 of the ewes received an oral dose of 3 mg melatonin in a 4:1 (v:v) mixture of water and ethanol daily at 15:00 h. The remaining 16 ewes received the vehicle alone. Within each of these groups, eight were implanted s.c. on day 0 with an osmotic minipump which infused 50 micrograms of the gonadotrophin releasing hormone agonist (GnRHa), buserelin day-1. On day 25, a second minipump was inserted to ensure continued infusion of the agonist and on day 50 (24 June) both minipumps were removed. Ovarian activity was assessed by laparoscopy at intervals of 3 weeks from day 29 until the experiment was terminated on day 200 (21 November). Blood samples taken by jugular venepuncture three times a week for the first 50 days, daily from days 51-78 and thereafter three times a week were analysed for progesterone, prolactin and LH. Samples taken at intervals of 15 min for 10 h on days--1, 14, 28, 49, 56, 70 and 91 were assayed for LH. Treatment with GnRHa reduced LH concentrations and abolished pulsatile LH secretion.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Neutrophil function in whole blood and after purification: changes in receptor expression, oxidase activity and responsiveness to cytokines.

Neutrophil function and plasma membrane receptor expression was measured in cell suspensions isolated by two separate procedures and in unfractionated whole blood. When cells were prepared by a combined dextran/ficoll procedure, their ability to generate reactive oxidants in response to fMet-Leu-Phe was greater than in corresponding cells isolated by a one-step procedure on Mono-Poly Resolving Medium (M-PRM). Cells prepared by both methods could be primed in vitro by rGM-CSF, but the priming ratio was greater in cells prepared by the latter method. The ability of neutrophils in whole blood to generate reactive oxidants in response to fMet-Leu-Phe was extremely low, but this was increased by more than 10 fold if the blood was pre-incubated with rGM-CSF. Similarly, expression of CD 11b and CD 16 was very low (or undetectable) in neutrophils in whole blood, but this was rapidly increased upon priming. Activation by PMA resulted in a down regulation of CD 16 expression as the receptor was shed from the cell surface. Neutrophils isolated by either the dextran/ficoll or the M-PRM method showed increased expression of receptors compared with those in whole blood, although this expression was lower in cells isolated by the latter method. These data indicate that the isolation procedures used to obtain purified neutrophils prime both receptor expression and oxidase function, although these effects are minimalised in isolation procedures using M-PRM. Furthermore, as CD 16 expression on neutrophils in whole blood is rapidly up-regulated during priming, it seems likely that, as for complement receptors, rapidly-mobilisable intracellular stores of this receptor exist.

Antibodies, Monoclonal↗
PubMed1992

Hyalin, a sea urchin extraembryonic matrix protein: relationship between calcium binding and hyalin gelation.

The protein hyalin, a major component of the sea urchin extraembryonic hyaline layer, was previously shown to undergo a Ca(2+)-induced self-association into large aggregates (gelation). This reaction represented a major step in assembly of the layer. In the experiments reported here, digestion with trypsin resulted in a rapid dissociation of hyalin into a mixture of peptides which retained the capacity to bind Ca2+. However, unlike intact hyalin, none of these peptides associated into large aggregates (gelation) in the presence of Ca2+, Mg2+, and NaCl. Loss of the ability to undergo gelation was not accompanied by any significant change in the content of acidic plus amide amino acid residues. Decreasing the pH to 5.6 resulted in a loss of 25% of hyalin's Ca(2+)-binding capacity but had no effect on the ability of the protein to undergo gelation. Peptide fragments were only partially effective at inhibiting hyalin gelation. Clearly, not all the Ca(2+)-binding sites were required for hyalin gelation and Ca2+ binding alone was insufficient to drive this reaction. In addition, hyalin appeared to possess two classes of protein-protein interaction domains, one of which was essential for gelation.

Amino Acids↗
PubMed1992

Stimulation of neutrophils by insoluble immunoglobulin aggregates from synovial fluid of patients with rheumatoid arthritis.

Insoluble immunoglobulin aggregates present in the synovial fluid of patients with rheumatoid arthritis have been examined for their ability to activate reactive oxidant and granule enzyme secretion from bloodstream neutrophils. These insoluble complexes activated luminol chemiluminescence, but did not activate O2-, H2O2 or granule enzyme secretion and did not activate lucigenin chemiluminescence, which also measures reactive oxidant secretion. Hence, the luminol chemiluminescence detected after activation by insoluble immunoglobulin aggregates must be due to intracellularly generated reactive oxidants, i.e. produced within phagolysosomes. Because reactive oxidant and granule enzyme secretion has occurred within rheumatoid joints, other mechanisms of neutrophil activation must exist.

Arthritis, Rheumatoid↗
PubMed1992

Problems with paradigms in a caring profession.

The terms 'paradigms' and 'paradigm shifts' entered everyday language following the publication in 1962 of Kuhn's book The Structure of Scientific Revolutions. Kuhn's original usage of these terms in relation to 'normal' science has since been inappropriately applied to other areas of formal knowledge. Their use in relation to nursing is conceptually flawed, and is an unnecessary distraction from the more modest, but nevertheless rigorous, approaches to the elucidation and generation of knowledge for practice which are required.

Humans↗
PubMed1992

Sequential phospholipase activation in the stimulation of the neutrophil NADPH oxidase.

Stimulation of human neutrophils with the chemotactic peptide fMet-Leu-Phe results in activation of a rapid, transient burst of oxidant secretion, which reaches a maximal rate by about 1 min after stimulation. This phase of oxidant secretion is then followed by intracellular oxidant production, which is detected by luminol chemiluminescence but not by assays such as cytochrome c reduction or scopoletin oxidation. The rapid phase of oxidant secretion requires increases in intracellular free Ca2+ and phospholipase A2 activity, but not the activities of phospholipase D or protein kinase C. In contrast, intracellular oxidant production requires the activities of phospholipase D and protein kinase C. A model is thus proposed suggesting the sequential activation of different phospholipases which activate oxidase molecules on the plasma membrane or else from the membranes of specific granules.

Calcium↗